Project Summary/Abstract: Head and neck squamous cell carcinoma is the sixth most common malignancy worldwide. Metastasis reduces survival rates for patients to only 37%, indicating the severity of advanced disease, but there remains much to be understood about the metastatic process. Emerging evidence suggests the tumor microenvironment contributes to HNSCC progression and this proposal aims to clarify the role of one microenvironment cell type, carcinoma-associated fibroblasts (CAFs). HNSCC has a high propensity for metastasis to the lymph nodes, and correlative studies have indicated high expression of the transmembrane glycoprotein podoplanin (PDPN) is indicative of poor patient prognosis. It has been suggested that CAFs may be expressing PDPN in the lymph nodes, though this has yet to be determined. In addition, PDPN+ CAFs in other cancer types have been shown to have varying roles in either promoting or inhibiting cancer progression though little data exists for HNSCC. This project will expand upon my preliminary data indicating CAFs from both the primary tumor and tumor-associated lymph nodes are, indeed, PDPN+ and accelerate HNSCC migration in vitro. The overarching objectives are to identify the mechanism of PDPN+ CAFs in facilitating HNSCC invasive and migratory potential, as metrics of metastatic potential, as well as assess the ability of the PDPN+ CAFs to enhance lymphangiogenesis and to seed a pre-metastatic, immunosuppressed, tumor-supportive niche in the lymph nodes. In vitro analyses will investigate the PDPN-dependent secreted factors that may influence HNSCC progression as well as downstream signaling of PDPN within the CAFs that may result in increased cell contractility and immunosuppressive abilities. All in vitro data will be complemented by mouse studies in the presence and absence of PDPN expression with outcomes analyzing lymphatic vasculature density, lymph node immune infiltration, and overall metastatic tumor burden. Study findings are expected to identify PDPN and/or its effectors as (a) potential target(s) for the treatment of metastatic HNSCC in an effort to reduce mortality rates caused by the devastation of metastatic occurrence.